In an effort to understand the nature of the interaction between a hormone and its target cell, it is important to localize and characterize the hormone receptor. This project is devoted to the systematic morphological and biochemical analyses of the prolactin receptor site(s) in/on rat luteinizing granulosa cells. To this end, "plated" and cultured granulosa cells will be incubated first with 125I-prolactin under varying conditions of differentiation, temperature, time, pH and various concentrations of labeled and excess unlabeled prolactin to determine optimal conditions for high-affinity and specific binding to prolactin receptors. Once these conditions have been determined, granulosa cells, subcellular fractions and frozen and epon sections of cells and cellular fractions will be incubated with 125I-labeled or ferritin labeled prolactin or with labeled antiserum directed against prolactin or its antiserum in order to localize the distribution pattern of prolactin receptor(s) in/on cells and cell fractions. High resolution autoradiography, cytochemical and immunocytochemical techniques will be employed. In other experiments, granulosa cells will be incubated with labeled prolactin or its labeled antiserum and at timed intervals thereafter subfractionated and analyzed for the presence of the labeled hormone. The degree of binding will be correlated with the capacity of the labeled hormone to promote morphological differentiation of mitochondria and progesterone secretion by these cells. This investigation will represent the first such study of the direct morphological localization of 125I-and ferritin-labeled prolactin in/on isolated ovarian cells and the correlation of this specific binding with the capacity of eliciting a biological response.